The demonstration of kinetic evidence for active monomers of muscle creatine kinase, a dimeric enzyme, poses the question of whether this extends to other isozymic forms as well as the physiological significance of subunit association. We propose to subject the brain type isozyme from monkey to extensive analysis including the denatured and renatured states and the kinetics of renaturation, reactivation and reassociation. We plan also to further characterize muscle creatine kinase to ascertain the dependence of activity and tertiary structure on the state of subunit interaction. Furthermore, synergism in substrate binding and formation of the transition state analog will be studied as they relate to quaternary structure. Among the techniques employed will be kinetic analysis using the pH stat assay, fluorescence studies involving corrected spectra and polarization and susceptibility to proteolytic digestion. A series of experiments are proposed, using a cross-linked dimer, to distinguish between renaturation and reassociation, to determine the kinetics of reassociation and to measure the dependency of regulatory properties on subunit interactions. In view of the demonstrated occurrence of mutated forms of creatine kinase in certain myopathies, we believe that an understanding of the relationship between subunit association, activity and regulation of the isozymes will clarify the possible role of creatine kinase in neuromuscular dysfunction.